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P450 Enzymes
- 1st Edition, Volume 693 - November 13, 2023
- Editor: Hans Renata
- Language: English
- Hardback ISBN:9 7 8 - 0 - 4 4 3 - 1 3 6 9 5 - 5
- eBook ISBN:9 7 8 - 0 - 4 4 3 - 1 3 6 9 6 - 2
P450 Enzymes, Volume 692 in the Methods in Enzymology series, highlights new advances in the field with this new volume presenting interesting chapters written by an international… Read more
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Request a sales quoteP450 Enzymes, Volume 692 in the Methods in Enzymology series, highlights new advances in the field with this new volume presenting interesting chapters written by an international board of authors. Chapters in this new release cover Non-natural P450 reaction (carbene/nitrene), Non-natural P450 reaction (atom transfer radical cyclization), P450 fusion for biocatalysis, P450 fingerprinting for biocatalytic oxidation, Fungal peroxygenase, Fungal peroxygenase, Anti-Markovnikov P450 oxygenase (AmOX), Fungal P450, Unusual P450 chemistry from fungal sources, CASTing for P450 biocatalysis, and P450 in C–C coupling of cyclic dipeptides with nucleobases.
- Provides the authority and expertise of leading contributors from an international board of authors
- Presents the latest release in the Methods in Enzymology serials
- Updated release includes the latest information on P450 Enzymes
Biochemists, biophysicists, molecular biologists, analytical chemists, and physiologists
- Cover image
- Title page
- Table of Contents
- Series Page
- Copyright
- Contributors
- Preface
- Chapter One: Directed evolution of P411 enzymes for amination of inert C–H bonds
- Abstract
- 1 Introduction
- 2 Generation of mutant libraries
- 3 Protein expression
- 4 Enzymatic reaction setup
- 5 Screening and validation of the enzymatic reactions
- 6 Summary
- References
- Chapter Two: P450-catalyzed atom transfer radical cyclization
- Abstract
- 1 Introduction
- 2 Materials
- 3 Protocols
- 4 Summary
- Acknowledgments
- References
- Chapter Three: Self-sufficient P450-reductase chimeras for biocatalysis
- Abstract
- 1 Introduction
- 2 Protocols
- 3 Biocatalytic reactions with PtmO5-RhFRed
- 4 Developing TleB as a self-sufficient P450 biocatalyst
- 5 Biocatalytic reaction with TleB-RhFRed
- Acknowledgments
- References
- Chapter Four: The colors of peroxygenase activity: Colorimetric high-throughput screening assays for directed evolution
- Abstract
- 1 Introduction
- 2 Materials, equipment and reagents
- 3 Enzyme production
- 4 Validation of the assay
- 5 High-throughput screening (HTS) assays
- 6 Discussion and outlook
- References
- Chapter Five: Engineering cytochrome P450s for selective alkene to carbonyl oxidation
- Abstract
- 1 Introduction
- 2 General methods
- 3 Library construction
- 4 Recombinant protein expression in 96-well plate format
- 5 Library screening
- 6 Rescreening of variants containing beneficial mutations
- 7 Summary and conclusions
- Acknowledgment
- References
- Chapter Six: Bacterial cytochrome P450 enzymes: Semi-rational design and screening of mutant libraries in recombinant Escherichia coli cells
- Abstract
- 1 Introduction
- 2 Cloning of surrogate redox partners for CYP107Z
- 3 Construction of focused P450 mutant library
- 4 Expression of site-saturation libraries and whole-cell biotransformation of (−)-deoxypodophyllotoxin in 96-deep-well plates
- 5 MISER LC/MS analysis
- 6 Verification of selected P450 variants for in vitro conversion of (−)-deoxypodophyllotoxin
- 7 Summary and conclusion
- References
- Chapter Seven: Functional analysis of a fungal P450 enzyme
- Abstract
- 1 Introduction
- 2 In vivo analysis of Trt6 and Trt14
- 3 In vitro analyses of Trt6 and Trt14
- Acknowledgments
- References
- Chapter Eight: Protein engineering using mutability landscapes: Controlling site-selectivity of P450-catalyzed steroid hydroxylation
- Abstract
- 1 Introduction
- 2 Mutability landscapes
- 3 Identification of hotspots by mutability landscape and empirical mutations
- 4 Enzyme engineering by iterative saturation mutation (ISM) and rational design
- 5 Exploration of the mutants to hydroxylate other steroids
- 6 Materials, equipment and reagents
- 7 Protocol
- 8 Summary
- Acknowledgments
- References
- Chapter Nine: P450 in C–C coupling of cyclodipeptides with nucleobases
- Abstract
- 1 Introduction
- 2 General instruction and cultivation of bacterial strains
- 3 Genomic DNA isolation
- 4 PCR amplification for P450 gene cloning
- 5 Cloning of plasmids and transformation of E. coli
- 6 Heterologous gene expression in Streptomyces
- 7 Protein overproduction in E. coli and purification
- 8 SDS-PAGE
- 9 Enzyme assays
- 10 Product isolation and structure elucidation
- 11 Summary and conclusions
- Acknowledgments
- References
- Chapter Ten: Secretion and directed evolution of unspecific peroxygenases in S. cerevisiae
- Abstract
- 1 Introduction
- 2 Generation of a signal peptide library
- 3 Library cultivation
- 4 Colorimetric screening methods
- 5 Isolation of episomal plasmids from S. cerevisiae
- 6 Golden Mutagenesis
- 7 Performing reactions in 96-well microtiter plate
- 8 MISER-GC–MS
- 9 Summary and conclusion
- Acknowledgment
- References
- Chapter Eleven: Direct aromatic nitration by bacterial P450 enzymes
- Abstract
- 1 Introduction
- 2 Creation of self-sufficient TxtE chimeras
- 3 Spectral analysis of TxtE fusion variants
- 4 Evaluation of TxtEBM3R chimera nitration activity
- 5 Summary and conclusions
- Acknowledgments
- References
- Chapter Twelve: P450 fatty acid decarboxylase
- Abstract
- 1 Introduction
- 2 Discovery of P450 fatty acid decarboxylases
- 3 Expression, characterization and structure analysis of P450 fatty acid decarboxylases
- 4 Engineering of P450 fatty acid decarboxylases
- 5 Applications of P450 fatty acid decarboxylases
- 6 Conclusion
- Acknowledgments
- References
- Chapter Thirteen: Directed evolution for Si–C bond cleavage of volatile siloxanes in glass bioreactors
- Abstract
- 1 Introduction
- 2 Generation and characterization of enzyme libraries
- 3 Assembling the components for reactions and reaction analysis with the glass bioreactors
- 4 Screening enzyme mutant libraries in glass 96-well bioreactors
- 5 Modification of procedures for validating improved enzyme variants
- 6 Summary
- Acknowledgements
- References
- No. of pages: 424
- Language: English
- Edition: 1
- Volume: 693
- Published: November 13, 2023
- Imprint: Academic Press
- Hardback ISBN: 9780443136955
- eBook ISBN: 9780443136962
HR
Hans Renata
Prof. Hans Renata was born in Surabaya, Indonesia. After completing his high school education in Singapore, he moved to the US and received his B.A. degree from Columbia University in 2008, conducting research under the tutelage of Professor Tristan H. Lambert. He earned his Ph.D. from The Scripps Research Institute in 2013 under the guidance of Professor Phil S. Baran. In his graduate thesis, Hans completed the synthesis of a polyhydroxylated cardiac glycoside, ouabagenin, and developed a range of corticosteroid analogues bearing hydroxylation at the C19 position. In 2013, he began his postdoctoral studies in the laboratory of Professor Frances H. Arnold at the California Institute of Technology. Hans joined the faculty at The Scripps Research Institute Florida in the summer of 2016 and was promoted to the rank of associate professor with tenure in 2022. Starting in July 2022, Hans will join the faculty at Rice University. Outside of the lab, Hans enjoys drinking good beer, while trying to stay up to date with new music and his favorite soccer teams
Affiliations and expertise
Associate Professor, Department of Chemistry, Rice University, Houston, TX, USA